In recent years, hosts for production of proteins of interest, such as animal cells (e.g., CHO), insects (e.g., silk worm), insect cells, animals (e.g., chickens and cows), and microorganisms (e.g., E. coli and yeast), have been used in order to produce proteins via genetic recombination. In particular, yeast cells can be cultured in large-scale, high-density culture systems in cost-effective media, and proteins can be produced at low cost. In addition, proteins can be secreted and expressed in a culture solution by using a signal peptide or the like, and a process for purifying proteins thus becomes easy. Further, yeast cells are advantageous in that they are eukaryotic organisms and post-translational modification such as glycosylation is possible. Accordingly, various studies have been made on yeast cells.
For example, Non-Patent Document 1 discloses a method for producing the green fluorescent protein, human serum albumin, the hepatitis B surface antigen, the human epidermal growth factor, and hirudin with the use of methanol-assimilating yeasts.
Non-Patent Document 2 discloses a method of introducing a gene comprising a nucleotide sequence encoding a partial antibody ligated to a site downstream of the alcohol oxidase (AOX) promoter into a yeast cell, thereby producing a low-molecular-weight antibody.
In addition, Non-Patent Document 3 discloses that productivity of a target protein is improved by introducing a gene comprising a nucleotide sequence encoding a solubility tag (e.g., glutathione-S-transferase (GST) or maltose-binding protein (MBP)) ligated to a nucleotide sequence encoding a target protein into a yeast cell.
When a yeast is used as a host that produces a protein via gene recombination, various attempts have been made in order to improve the productivity. Examples of such attempts include the addition of a signal sequence or tag to a gene encoding a target protein, the use of a potent promoter, codon modification, chaperon gene co-expression, and examination of host culture conditions.